Instrumentation and Lab Equipment

Advanced Digital Microscopy

February 1, 2011
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Digital microscopes have been developed to overcome many conventional optical shortcomings.

With relatively few progressions in conventional optical microscopy during the past decade, pushing through the physical limitations of these systems has proven to be challenging. Despite this stand-still, engineers have recently been able to develop solutions to some of the limitations in modern microanalysis.

Using an integrated design approach, digital microscopes have been developed to overcome many of the shortcomings of conventional optical microscopes. One of the major difficulties in observing and analyzing ceramic materials is their typical complete lack of contrast, especially when viewed through a traditional optical microscope. However, the advanced features of fully integrated digital microscopes, coupled with high-resolution zoom lenses and digital camera technology, can enable users to image samples that were previously relegated to the realm of a scanning electron microscope (SEM).

Figure 1. Advanced digital microscope systems can produce an extended depth-of-field image in real time.

Increasing the Depth of Field

Relatively few technological advances have been made from decades-old optical microscopes to those on the market today. Conventional microscope systems have virtually reached their peak in standard configuration and optical design, with emphasis on the reduction of chromatic and spherical aberration of the optics. High-resolution and high-magnification observation appeared to be achievable with the advancements in zoom lens optics, digital camera and imaging technologies.

For conventional optical technologies, however, depth of field was lost as the magnification increased or when a digital camera was attached to the microscope system, causing higher magnification image capture to become problematic. With the development of integrated digital microscopes over 20 years ago, it was thought that future improvements in electronics and semiconductor technology would help facilitate the advancement of microscopy.

One of the major obstacles in material inspection applications is the limited depth of field offered by conventional microscope optics. Typical microscope objectives can only focus on a small section of a target at a given time. Optimizing the optical properties of a lens to increase its working distance and expand its depth of field while still producing a high-resolution image was a major challenge facing digital microscope developers. While stereoscopes have the ability to achieve this type of performance, the depth of field is lost when a camera is attached. The optimally designed optics and integrated charge-coupled device (CCD) camera of a digital microscope work together to capture sharply focused images with a high depth of field. This enabled a dramatically increased ability to observe surface structure and detail.

Naturally, when the magnification is increased, the depth of field is diminished. For higher magnification observation, a function was developed to produce an extended depth-of-field image (see Figure 1). While some systems must capture and then process the images on a separate PC, more advanced digital microscope systems can produce an extended depth-of-field image in real-time through an adjustment to the focus knob. As the lens is moved through the different focal planes, an algorithm can dynamically capture only the focused pixels, compiling them into a fully focused image. Due to the brittle nature of ceramics, the observation of fractured areas is a common application that is not easily accomplished with typical optical systems.

This same algorithm can also be used to produce a 3-D model of the surface of an object (see Figure 2). The relative heights at which pixels come into focus are stored and then used to reconstruct the surface topography of the target. 3-D measurements such as volume, distance, angle and others can be obtained with an accuracy of +/-1 micron at 1000x magnification. After a 3-D model of surface topography is generated, profile information such as depth can be generated and exported as a CSV file.

Figure 2. 3-D image reconstruction of a ceramic surface. (Lighting adjustment used to enhance surface texture.)

Free-Angle Viewing

When using a conventional industrial microscope, metallurgical scope or stereoscope, the user is presented with the challenge of perfectly mounting their sample for observation. Most industrial inspection applications require multi-angle viewing. Typically, the user needs to either mount the part with putty or even hold that part in their hands to observe all of the sample's surfaces.

Advanced digital microscope systems can be equipped with a multi-angle stand, allowing the user to tilt the lens and camera around the target being viewed. The lens is mounted into the arm of the stand, which is equipped with a coarse and fine adjustment focusing knob. This arm can also be upgraded to precision motorized control.

The benefit of this multi-angle viewing stand is that the sample simply needs to be placed onto the stage for viewing. Instead of manipulating the target itself, the lens is actually manipulated around the target, providing almost 180° rotation about the Z axis. In addition, the X-Y stage rotates 360° to allow for complete multi-angle viewing of the part without any mounting or hand manipulation. This is particularly useful when viewing a fractured surface, since a top-down view will not typically provide the user with all of the necessary information regarding the cause of the failure.

Figure 3. Interpolation vs. pixel-shift method.

Resolution and the Pixel-Shift Method

Whether for basic observation or image capture, a higher resolution image generally provides for better analysis. Ceramic materials can have extremely detailed surfaces, often requiring a high-resolution image in order to adequately capture all of the unique characteristics of a particular sample. Since more advanced digital microscope systems are completely integrated, the resolution and color clarity of the camera is integral to the performance of the system.

Typical camera systems used for microscopy have a basic CCD resolution of about 1.5-5 megapixels. In order to provide users with a larger image file, techniques are used with these systems to increase the resolution of the resulting image. Interpolation is a common method, whereby the jagged, pixilated edges of an image are smoothed using an average of pixel coloration along the edges of a target. When the user zooms in on the resultant image, this method effectively reduces the jaggedness of the pixilated edges. A disadvantage of this technique, however, is that the resolution of the image is not actually increased since pixel information is interpolated.

Instead of interpolating data, higher end systems use the pixel-shift method to increase the resolution of a capture image (see Figure 3). Piezo-crystal actuators are attached to the different sides of the CCD. When supplied with a voltage, these crystals expand and physically shift the CCD, ultimately shifting it in a 3X3 array, capturing pictures at each new position. This process creates nine real sub-pixels for every pixel, increasing the pixel value of the CCD image from 2 megapixels to 18 megapixels, for example.

Figure 4. Operating principle of the HDR function.

Lighting Optimization

Viewing a sample under optimal lighting can make the difference between a mediocre image and a groundbreaking one. Different types of lighting accentuate various features on a sample; in certain lighting, some of these features may go unseen.

One of the major challenges of traditional optical microscope systems is their inability to create different reproducible lighting scenarios for observation. Digital microscopes with zoom lenses can integrate the lighting with the lens so the target in the field of view is always viewed under optimal lighting conditions. For highly reflective samples or other lighting challenges, a full lineup of lighting adapters for the lenses (e.g., diffuse, polarized and variable-angle lighting) can be used.

Several different software functions can also be employed to push beyond these limits to obtain a sample's optimum brightness and contrast. From there, the user can choose between combinations of edge, color or contrast enhancing illumination to quickly find the most suitable lighting for the application. Even when the lighting scenario has been chosen, the user can switch and apply different configurations to the image.

Figure 5. White ceramic surface shown in conventional (here) and HDR images (below).

One of the most distinctive and useful of these reproducible lighting scenarios is the High Dynamic Range (HDR) function (see Figure 4). The HDR function first captures multiple images at different brightness levels, and then compiles them into a single image with exponentially higher levels of color gradation. The HDR function takes the image from an 8-bit color resolution (256 levels of color gradation) to 16-bit color resolution (65,536 levels of color gradation), allowing the user to see details on the target that weren't observable with conventional 8-bit imaging.

Specifically, the HDR function generates images with much higher contrast than conventional microscope images, produces vivid color and also eliminates glare from reflective surfaces (see Figure 5). Furthermore, the increased image data (e.g., texture, brightness, color and contrast) can be amplified or suppressed with a slide-bar to isolate or accentuate different features of the sample. By suppressing the texture, for example, users can isolate particles of a specific color without having shadows affect the color extraction process.

Figure 5

Precision and Versatility

In order to make measurements using a conventional optical microscope system, users must capture the image, export it to a PC, and use separate analysis software to obtain any measurements. Digital microscopes integrate both 2- and 3-D measurement capabilities. Users can make 2-D measurements like radius, distance, angle, or area on a 2-D image, while 3-D measurements such as volume, angle, distance, or profile are also possible. Individuals concerned with the quality of their ceramics can easily measure features like micro-cracks or porosity and reduce the amount of time needed for inspection (see Figure 6).

Figure 6. 2-D measurement and documentation.

While conventional optical microscope magnifications are specified based on the product of the objective and ocular lenses, the definition for digital microscope magnification is quite different. Since these systems usually employ zoom-style tube lenses to display and image directly on a monitor, the magnification is calculated based on the size of the medium used to view the target. In most cases, all-in-one digital microscopes display an image on a 15-in. screen and define their magnifications accordingly. For example, a typical magnification range for an integrated digital microscope is 0.1-5000x.

In addition to their high resolution and large depth-of-field, another major benefit of these lenses is their working distance. Conventional optics, while providing a high-resolution image, do not usually allow a user much space between the top of the sample and the bottom of the objective lens. At 1000x, for example, the working distance for an objective lens can be as small as 0.2 mm. Zoom lenses can achieve a similar magnification while still maintaining a working distance of over 4 mm, with some lenses providing as much as 4 in. or more at other magnifications.

As scientists find increasing applications for ceramics, the need for versatile lenses will become a necessity for inspection and development work. By using a digital microscope with zoom lens, applications in ceramics such as examining disk brakes, turbine blades, medical implants, ball bearings and armored vests can be easily performed.

For more information, contact KEYENCE Corp. at 1100 North Arlington Heights Rd., Itasca, IL 60143; (888) 539-3623; e-mail; or visit


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